UC Davis researchers have affirmed an essential hypothesis in a 26-year-old DNA repair model – new evidence paving the way for further study into the molecular mechanisms that can affect cancer predisposition and developmental defects.
Before this work, the 1983 double-strand-break repair model’s key hypothesis had not been physically demonstrated in cells, said Wolf-Dietrich Heyer, professor of microbiology.
“This research is a really big step,” Heyer said.
Intermediate DNA structures called double-Holliday junctions are formed in vivo when the ends of a broken chromosome – a single piece of coiled DNA and protein found in cells – exchange strands with a second chromosome, according to a paper by department of microbiology research scientist Malgosia Bzymek. The paper is published in the Apr. 8 issue in the journal Nature.
“In this repair process called homologous recombination, the second chromosome acts as a template from which the broken chromosome can copy any missing or damaged DNA sequences,” said Neil Hunter, associate professor of microbiology and senior author on the paper, in an e-mail interview.
The work also confirmed that double-Holliday junctions appear in both the cell divisions of meiosis and mitosis, an occurrence previously surmised by researchers, but scientifically undetermined, Heyer said.
Throughout meiosis, or sexual reproduction, cells divide twice to produce sperm and egg cells.
“Crossovers are at the heart of this process; without the connections they provide, mom and dad chromosomes are often pulled in the same direction, and the resulting sperm or eggs have odd numbers of chromosomes,” Hunter said in a statement.
Throughout mitosis – the cell division process in which the body divides its non-sexual cells – a cell identically splits into two. But before the cell can divide, its chromosomes must be entirely copied. And homologous recombination helps fix breaks that arise during that DNA replication, according to a UC Davis press release.
“Homologous recombination is an essential repair process that helps fix problems that frequently occur as cells replicate their chromosomes,” Hunter said. “But homologous recombination is something of a double-edged sword because aberrations in the process cause chromosomal changes that can lead to cancer.”
Double-Holliday junctions can also resolve into crossovers – entire chromosome arms are exchanged at the site of repair.
“Crossovers can cause genetic changes that are tumorigenic,” Hunter said. “We also discovered that double-Holliday junctions represent a minor pathway of break repair, i.e. it appears that the cell actively prevents formation of double-Holliday junctions in order to limit crossovers and their associated problems.”
In the lab, the scientists devised an experiment to watch the repair happen at the molecular level. They observed the molecules breaking, interacting with the template chromosome, forming double-Holliday junctions and ultimately being restored.
Although the idea behind the research was not new, in many ways it represents a beginning, Hunter said.
“We’re especially interested to know how the formation of double-Holliday junctions is regulated.”
“This has really profound pragmatic value for future cancer research,” he said.
Co-authors of the study were Nathaniel Thayer, then a UC Davis undergraduate and now graduate student at the University of Washington; Steven Oh, then a UC Davis graduate student and now a postdoctoral fellow at UCSF; and Nancy Kleckner of Harvard University. The work was supported by the National Institutes of Health and the Howard Hughes Medical Institute, said a UC Davis press release.
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